Membrane for an in-vitro-resorption model of the gastrointestinal tract

ABSTRACT

A porous membrane adapted for use as the filter element in an in-vitro-resorption model of the gastrointestinal tract, said membrane consisting essentially of solid porous carrier material, especially porous cellulose nitrate, impregnated with a liquid phase consisting of a mixture of (1) a higher fatty acid, such as caprylic acid, or dioctyl sodium sulfosuccinate and (2) a neutral lipoid component, such as lauryl alcohol.

United States Patent 1191 Stricker 5] Aug. 21, 1973 MEMBRANE FOR ANIN-VITRO-RESORPTION MODEL OF THE [56] References Cited GASTROINTESTINALTRACT UNITED STATES PATENTS Inventor: Herbert Stricken Ingelheim am3,612,283 10/1971 Stricker 210/490 Rhein, Germany [73] Assignee: C. H.Bolhringer Sohn, Ingelheim Primary Examiner-Frank A. Spear, Jr.

am Rhein, Germany Att0rneyHammond & Littell [22] Filed: July 27, 1971 57ABSTRACT [21] Appl. No.: 166,350 1 A porous membrane adapted for use asthe filter ele- Related Application Data ment in an in-vitro-resorptionmodel of the gastrointes- [63] Continuation-impart of Ser. No. 861,777,Sept. 29, tinal tract, said membrane consisting essentially of solid1969, Pat. No. 3,612,283. porous carrier material, especially porouscellulose nitrate, impregnated with a liquid phase consisting of a [30]Foreign Application Priority Data mixture of (1) a higher fatty acid,such as caprylic acid,

Sept. 27, 1968 Germany P 17 98 340.3 or dioctyl sodium sulfosuccinateand a neutral poid component, such as lauryl alcohol. [52] U.S. Cl.210/490, 210/506 511 rm. c1 B0ld 39/14, 801d 31/00 2 Chums 2 F'gures[58] Field of Search 210/23, 321, 490,

BACKGROUND OF THE INVENTION The effectiveness of a drug in the system.of the human body is, inter alia, a function of its concentration atthe place of effective action and thus also a function of its ability toreach that place. Most of the perorally administered drugs must firstovercome the barriers of the gastrointestinal tract, i.e., they must beresorbed, before they can be transported by the blood to the place wherethey exert their therapeutic action.

Various in-vivo methods for following'the passage of chemo-therapeuticsfrom the gastrointestinal tract into the blood circulation system areknown.

For instance, the decrease in the concentration of the chemo-therapeuticsubstance with passage of time in various parts of the gastrointestinaltract after peroral administration can be determined by analyticalmethods in the tied-ofi stomach, colon or small intestine of rats.

In humans, the change in concentration of various dissolved organicacids and bases in the stomach over a period of time can also befollowed by means of an esophageal sound lowered into the stomach.

Likewise, in humans as well as animals, the blood level values atvarious times after perorjal administration of drugs can be determined,from which rate of resorption can then be calculated.

Such in-vivo methods, however, require a substantial expenditure of workand money, and the results obtained therefrom are often subject to widevariations with respect to accuracy and reproduceability.

In-vitro methods employing artificial membranes, on the other hand,represent a valuable supplementation of the in-vivo determinations.Thus, by means of greater series tests and more accurate concentrationdeterminations, the kinetics of diffusion can be very accuratelyascertained, so that the effects of the interchange action between theactive ingredients among themselves or with the pharmaceuticalexcipients can already be evaluated and taken into consideration indetermining the most favorable make-up of the pharmaceutical dosage unitcompositions. Moreover, by employing in-vitro methods, the number ofin-vivo tests can be significantly reduced.

THE PRIOR ART Filter membranes for various purposes have already beendescribed in the literature. For instance, porous membranes made ofcellulose esters and impregnated with olive oil or linoleic acid havebeen described by G. Levy and E. .l. Mroszezak in J. Pharm.Sci. 57(2),235-239 (1968).

However, it was found that, while these olive oilimpregnated filtermembranes are pH-impermeable," they also do not permit the passage ofsubstances of an acid character if they are present in Weakly acid toneutral solution. Therefore, the olive oil'impregnated membranes are notwell suited for in-vitro resorption tests, especially if the apparatusis to simulate resorption in the small intestine.

On the other hand, linoleic acid-impregnated membranes proved to beinsufficiently permeable with respect to hydrogen or hydroxyl ions, sothat they could not be used for diffusion measurements between solutionshaving a different pH-value.

OBJECTS OF THE INVENTION The present invention has as its principalobject to provide a membrane for use in an in-vitro resorption testapparatus, which is capable of maintaining the pH- gradient between twoaqueous solutions substantially constant over a period of time of about50 hours and exhibits a specific permeability for acid, basic andamphoteric drug molecules.

Another object of the instant invention is to provide a porous membranefor use in an in-vitro resorption test apparatus, which duplicates asclosely as possible the drug diffusion rates under physiologicalconditions known from in-vivo tests.

Other objects and advantages of the invention will become apparent asthe description thereof proceeds.

THE INVENTION The above objects are achieved in accordance with thepresent invention by impregnating a solid porous carrier material with aliquid lipoid phase.

The porous carrier material may be a commercial membrane filter materialmade of a cellulose ester. The diameter of the pores must besufficiently small so that the pores, due to their capillary activity,remain closed by means of the liquid impregnation component; otherwise,the liquid impregnation phase could be dissolved out and the membranewould become useless. Therefore, the diameter of the pores should befrom 50 to 1,000 mu, preferably 100 mu. Examples of suitable membranefilter material are the commercially available products Millipore VC andSartorious SM 1 1309.

The liquid lipoid phase must resemble, with respect to its physicalproperties, such as its relative solution capacity for acid, basic andamphoteric drugs, and its distribution, as closely as possible to itsnatural counterpart, i.e. the stomach and intestinal walls, so that therelative resorption rates of various drugs can be accurately simulatedin the in-vitro apparatus.

I have found that these requirements are met by a mixture consisting ofl) a higher fatty acid, preferably caprylic acid, or dioctyl sodiumsulfosuccinate, and (2) a neutral lipoid component, such as a higherfatty alcohol of 1-2 or more carbon atoms, preferably lauryl alcohol.The mixture of these two components must be liquid at the testtemperature of 37C.

A membrane according to the present invention is prepared by immersingthe porous filter material into said liquid mixture, removing theimpregnated porous material from the liquid mixture and allowing excessliquid mixture to drip off, and blotting the impregnated porous membranebetween filter paper until its weight is constant. A preferredembodiment of the membrane is obtained by using as the liquidimpregnating component a mixture of parts by volume caprylic acid and 20parts by volume lauryl alcohol, or a mixture of 97 parts by volumelauryl alcohol and three parts by volume dioctyl sodium succinate.

In contrast to known membranes of this type, membranes in accordancewith the present invention have the advantage that they are capable ofmaintaining the pH-gradient between two aqueous solutions substantiallyconstant (i about 0.3 pI-l-units) over a period of about 50 hours andexhibit a specific permeability for acid, basic and amphoteric drugmolecules. The diffusion rates under physiological conditions correspondsubstantially to the values known from 'in-vivo tests.

The combination of a higher fatty acid or dioctyl sodium sulfosuccinateand a neutral lipoid component with a cellulose ester filter membrane asa carrier material proved to be a system which meets the requirementssurprisingly well. The effect of the fatty acid or sulfosuccinatecomponent is that, corresponding to its pK,-value and the pH of theadjacent solution, it is partly present as an anion and acts as aproton-acceptor or -donor with respect to the dissociated organic acidsor bases. The drugs are thereby transformed at the membrane surface intothe undissociated form and are thus capable of migrating through theliquid phase of the membrane, whereas the hydrogen and hydroxyl ions canpass through only very slowly because of the solubility conditions.

The annexed drawings show schematically the type of in-vitro diffusionapparatus wherein a membrane according to the present invention can beused.

FIG. 1 is a schematic representation of an apparatus which consistsessentially of two chambers l and 1a (volume of each 100 ml) which areseparated by a membrane 2 according to the present invention (effectivesurface 12.5 cm). Each of the chambers is provided with athermostatically controlled exterior heating jacket 3 to maintain thecontents at 37 i 0.5C. Two pipettes 4 and 5 extend into each of thechambers 1 and la, and a dropping funnel (not shown) is connected to theexterior end of each pipette. The flow of liquid through each pair ofpipettes is controlled by a double stopcock 6. Pipettes 5 terminatesubstantially at the point where they enter chambers 1 and la,respectively, whereas pipettes 4 extend into the respective chambers toa point close to the surface of membrane 2. One of the two chambers land 1a is filled with a physiological solution having a pH of 7.5, whichcorresponds to that of blood plasma; the other chamber is filled with aphysiological solution whose pH is adjusted to between 1.2 and 7.0,depending upon the section of the gastrointestinal tract which it isintended to duplicate. The dropping funnel connected to each of theshort pipettes 5 is filled with buffer solution to maintain the pH ineach chamber at the desired level. A metal disc 7 having a centralaperture is slidably mounted on each pipette 4. The entire apparatus ismounted so that it oscillates 36 times per minute through an angle of170 about a transverse axis 8 which coincides with the.

center of the plane formed by membrane 2, the oscillating motion beingprovided by a motor (not shown). By virtue of this oscillating motionthe metal discs 7 slide along pipettes 4 and thereby stir the solutionin each chamber. In order to withdraw a sample of the solution inchamber 1 or 1a, the respective double stopcock 6 is opened, whereby thedescending buffer solution in pipette 5 forces some solution from thechamber upward through pipette 4 into the dropping funnel connectedthereto, wherefrom it can be withdrawn for analysis. At the same timethe solution in the chamber is replenished with fresh buffer solution.The function of membrane 2 is to maintain the pI-I-gradient between thesolutions in chambers 1 and 1a, and also to permit differential passageof various drugs, i.e., organic acids and bases of different types. Thedrug whose diffusion rate is to be tested is introduced into thesolution which duplicates the particular section of the intestinaltract.

FIG. 2 is a schematic representation of another diffusion apparatuswhich comprises a diffusion vessel 9 consisting of two chambers 10 and10a separated by a membrane 11 according to the present invention. Thetest solutions are cycled by means of pump 12 from storage vessels 13and 130 into chambers 10 and 10a, respectively, and then back into thestorage vessels. The cycling rate depends on the dimensions of theapparatus; for example, if the membrane surface is about 12 cm and thevolume of chambers 10 and 10a is about 2 to 5 ml each, the cycling rateis advantageously about 2 to 10 ml/min. In addition, the solutions instorage vessels 13 and 13a are continuously stirred by means of amagnetic stirrer 14 in order to avoid concentration gradients.Thermostatically controlled heating jackets 15 maintain a constanttemperature of 37C. Stopcocks 16 permit withdrawal of samples of thesolutions for analysis.

The following examples further illustrate the present invention and willenable others skilled in the art to understand it more completely.

EXAMPLE 1 Using a diffusion test apparatus such as the one shown in FIG.1 of the drawings, the resorption rate of various acid and basic drugsfrom the stomach was tested.

The membrane was a porous sheet of cellulose nitrate, pore size i 8 mu(Millipore VC or Sartorius SM 11309), impregnated with a liquid mixtureconsisting of 50 parts by volume of caprylic acid and 50 parts by volumeof lauryl alcohol.

One diffusion chamber was filled with a solution of the drug compound in100 ml of a buffer solution having a pH of 1.2, i.e., the pH prevailingin the stomach, prepared from 24 gm of l N I-ICI, 0.5 gm of glycocoll,0.35 gm of NaCl and enough H O to make L000 ml.

The other diffusion chamber was filled with 100 m1 of a buffer solutionhaving a pH of 7.5, prepared from 20.5 gm of Na I-IPO 2.7 gm'of KH PQ,and enough H O to make 1,000 ml.

The apparatus was oscillated as described above in connection with FIG.1 of the drawings, and samples of the solution in each chamber werewithdrawn periodically, while at the same time replenishing the buffersolution from the dropping funnels.

The following table shows the results obtained in comparison to knownin-vivo resorption rates, all values being based on the resorption rateof aspirin 1.00

TABLEI Initial Resorption Rate, based on rate of acetylsalicylic acid1.00

* 1 S. Schanker et al .J. Pharm. exp. Therap. 120. 528(19581.

EXAMPLE 2 The test procedure and apparatus were the same as in Example1, except that the membrane was impregnated with a liquid mixtureconsisting of 20 parts by volume of lauryl alcohol and 80 parts byvolume of caprylic acid, and the buffer solution having the drugcompound dissolved therein had a pH of 6.0 to simulate conditions in thesmall intestine; the buffer solution was prepared from 1.5 gm of Na i-W80 gm of KH PO and enough water to make 1,000 ml.

The following results were obtained:

TABLEll Initial Resorption Rate, based on rate of acetylsalicylic acid1.00 Compound in-vivo' in-vitro Acetylsalicylic acid Benzoic acidBarbital Phenol Red Aminopyrine Ephedrine Theophylline Antipyrine L. S.Schanker et al., J. Pharm. exp. Therap. 120, 528 (1958).

EXAMPLE 3 TABLElll Initial Resorption Rate, based on rate ofacetylsalicylic acid 1.00 Compound in-vivo' in-vitro Acetylcalicylicacid 1.00 1.00 Salicylic acid 2.30 2.10 Benzoic acid 2.00 2.10 PhenolRed 005 0.05 Aminopyrine 0.05 0.02 Quinine 0.00 0.00 Ephedrine 0.00 0.00

* L. S. Schanker et al..J. Pharm. exp. Therap. 120. 528 (1958).

The results of these examples clearly show that, with the aid of anin-vitro apparatus comprising a membrane according to the presentinvention it is possible to duplicate very closely the in-vivoresorption conditions of the gastrointestinal tract.

While the present invention has been illustrated with the aid of certainspecific embodiments thereof, it will be readily apparent to othersskilled in the art that the invention is not limited to these particularembodiments, and that various changes and modifications may be madewithout departing from the spirit of the invention or the scope of theappended claims.

I claim:

1. A membrane for an in-vitro resorption model of the gastrointestinaltract, said membrane consisting of a porous cellulose ester filtermembrane material having a pore size of 0.05 to 1.0 mu impregnated witha mixture consisting of dioctyl sodium sulfo-succinate and laurylalcohol, said mixture being liquid at 37C.

2. A membrane according to claim 1, wherein said liquid mixture consistsof 3 percent by volume of dioctyl sodium succinate and 97 percent byvolume of lauryl alcohol.

2. A membrane according to claim 1, wherein said liquid mixture consistsof 3 percent by volume of dioctyl sodium succinate and 97 percent byvolume of lauryl alcohol.